Format

Send to

Choose Destination
See comment in PubMed Commons below
J Biol Chem. 1983 Oct 25;258(20):12644-53.

Activation of Na+/H+ exchange by epidermal growth factor elevates intracellular pH in A431 cells.

Abstract

Epidermal growth factor (EGF) increases Na+ uptake in several cell types through an electroneutral, amiloride-sensitive pathway putatively identified as Na+/H+ countertransport. We have previously shown (Rothenberg, P., Glaser, L., Schlesinger, P., and Cassel, D. (1983) J. Biol. Chem. 257, 4883-4889) that EGF rapidly activates amiloride-sensitive net Na+ influx in the A431 human epidermoid carcinoma cell line. We also described the presence of transmembrane, amiloride-sensitive Na+/H+ exchange in A431 cells using a new fluorescence technique for the measurement of intracellular pH (pHi) based on the incorporation of fluorescein-dextran into the cell cytoplasm. The low pKa of fluorescein (approximately 6.4) prevented the direct assessment of the inferred, EGF-induced cytoplasmic alkalinization, mediated by stimulated Na+/H+ exchange. In this paper, 4',5'-dimethylfluorescein (pKa 6.75) was coupled to dextran, allowing increased pH sensitivity of the fluorescence assay in the physiological range. Using this improved assay, basic features of pHi regulation in A431 cells are documented, including the role of Na+/H+ exchange and Na+-linked C1-/HCO3-exchange in acid extrusion. We directly demonstrate a rapid elevation of pHi by addition of EGF as well as by serum in A431 cells. The pHi increase is half-maximal at 5-10 ng/ml of EGF, is dependent on external Na+, independent of external Ca2+, and inhibited by millimolar amiloride. EGF and serum also enhance Na+/H+ exchange-mediated cytosolic acidification when the transmembrane Na+ concentration gradient favors Na+ efflux from the cells. An alkaline pHi shift, caused by activation of Na+/H+ exchange, may be an important primary event in the mechanism of EGF action.

PMID:
6195155
[Indexed for MEDLINE]
Free full text
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for HighWire
    Loading ...
    Support Center